The following discussion of the background of the invention is merely provided to aid the reader in understanding the invention and is not admitted to describe or constitute prior art to the present invention.
Stable storage for long periods of time typically requires cold temperatures. This is especially true for biological tissue. Water located between ice crystals contains concentrated solutes, and therefore remains a viscous fluid until the glass transition temperature is reached. At the glass transition temperature a crystal-free immobilized liquid state known as a “glass” is formed. This typically occurs between −80° C. and −130° C. Below the glass transition temperature, translational molecular motion and chemical conversions are arrested, and storage for indefinite periods of time becomes possible.
Frozen material is often stored submerged in liquid nitrogen (LN2) Dewars. LN2 has a temperature of −196° C. and is much colder than necessary for stable storage, but has the advantages of being inexpensive and readily available. Cold storage can also be accomplished in the cold vapor space that forms above a pool of LN2, which avoids concerns about contamination by LN2. Mechanical freezers operating between −80° C. and −150° C. are yet another method of cold storage.
Vitrification is an alternative to freezing for the preservation of biological material (e.g., see U.S. Pat. Nos. 4,559,298 and 5,217,860). In vitrification, water is replaced with one or more cryoprotective chemicals in order to completely suppress the formation of ice crystals during the drop in temperature. Unlike freezing, vitrification is not harmful to even the most complicated of living systems. Vitrification creates a “glass” state. The entire cell or tissue mass becomes a glass below the glass transition temperature, permitting storage at reduced temperatures for indefinite periods without structural alteration. Storage methods for vitrified cells or small tissue samples are the same as those for freezing.
The absence of structural damage makes vitrification attractive for cryopreservation of organs and other organized tissues. However vitrification of large objects involves problems that don't exist for small (microliter) objects (Fahy, et al., “Physical problems with the vitrification of large biological systems,” Cryobiology 27, pp. 492–510 (1990)). A major problem in vitrification is fracturing. Fracturing can occur if large objects are cooled far below the glass transition temperature, such as to liquid nitrogen temperature. Avoiding fracturing typically requires storing within 10° C. to 20° C. of the glass transition temperature. The possibility of fracturing is also reduced if an annealing process is performed in which the temperature approaches the final storage temperature sigmoidally, and very slowly (Baudot, et al., “Physical Vitrification of Rabbit Aortas without any Fracture,” Cryobiology 43, p. 375 (2001)).
Presently available storage systems are designed to accommodate many objects, which are placed in a single isothermal or poorly-controlled vapor environment. These systems are not suitable for banking of large tissue masses where fracturing is a concern. Furthermore, annealing protocols require the storage temperature to be manipulated over potentially long periods of time. Different objects must be held at different temperatures, depending on the phase of their annealing process. Different objects may even have different final storage temperatures because the glass transition temperature can differ depending on the cryoprotectant mixture used to treat the object.
Accordingly, a need exists for devices that can store materials at cryogenic temperatures in a controlled temperature environment.